Release of FSH from the anterior pituitary is essential for gametogenesis, as FSH results in stimulation of spermatogenesis in the testis, as well as maturation of the oocyte and secretion of estradiol by the ovarian follicles. The biosynthesis and secretion of FSH within the anterior pituitary is controlled through the complex interaction of several hormones. These include hypothalamic gonadotropin-releasing hormone (GnRH), gonadal steroids, and the recently-identified gonadal peptides, inhibin, activin, and follistatin.
Inhibin and follistatin have the ability to inhibit the synthesis and secretion of FSH by the pituitary (De Jong et al., Nature 263: 71-72 (1976)), while activin has been shown to be capable of stimulating FSH synthesis and release (Vale et al., Nature 321: 776-779 (1986); Ling et al., Nature 321: 779-782 (1986)). Inhibin is a heterodimeric glycoprotein composed of an .alpha. subunit linked to one of two .beta. subunits (.beta..sub.A or .beta..sub.B), whereas activin consists of heterodimers or homodimers of inhibin .beta. subunits. Although originally isolated from the gonads, both inhibin and activin have been detected in the pituitary where they play an autocrine/paracrine role in the control of FSH gene expression (Meunier et al., Proceedings of the National Academy of Sciences 85:247-251, (1988)). In addition, recent immunohistochemical studies have determined that inhibin/activin subunit proteins localized in the pituitary gonadotropes are responsible for FSH and LH synthesis (Roberts et al., 1989, Endocrinology 124:552-554).
Both activin and inhibin subunits have been isolated from natural sources. In Mason et al., U.S. Pat. No. 4,798,885, the amino acid sequences of the .alpha., .beta..sub.A and .beta..sub.B subunits of human and porcine inhibin are described and are used to identify the amino acid sequences of activin. Mason et al. discuss the use of activin as a fertility-inducing therapeutic capable of stimulating FSH release.
The use of activin to increase fertility in a male mammal is described in Attie et al., PCT No. 91/10444. Following treatment of rat testicular germ cells with activin, these cells were observed to proliferate 2.0-2.5 times faster than control cells. Additional studies have confirmed the initial description of activin's ability to stimulate FSH secretion from cultured rat pituitary cells by about 2-3 fold (Vale et al., Nature 321: 776-779 (1986); Ling et al., Nature 321: 779-782 (1986); Carroll et al., Molecular Endocrinology, 3:1969-1976 (1989); Kowaga et al., Endocrinology 128, 1682-1684 (1991); Attardi et al., Mol. Endo. 4: 721 (1990)). Although these results raise the possibility that activin may increase fertility in mammalian subjects, the measured increase in FSH secretion is typically small.
Recent results suggest that the importance of activin to maintenance of FSH biosynthesis is much greater than has previously been recognized. The mRNA for the .beta. subunits of FSH become rapidly undetectable in pituitary cells cultured in a way to remove endogenous-secreted activin (Weiss et al., Endocrinology 131: 1403-1408 (1992)). When activin is then added back to the medium, these low levels of FSH mRNA increase 30-70 fold, a substantially greater increase than the 2-3 fold stimulation that is observed when cells are cultured in the presence of endogenous activin.
The biological activity of FSH can be influenced by the presence of a binding protein, such as follistatin (FS) (Ling et al., Nature 321: 779-782 (1986)), which may form complexes with activin (Krummin et al., Endocrinology 132: 431 (1993)). Recent studies indicate that incubation of rat pituitary cultures with an immunoneutralizing antibody raised against activin results in a decline in FSH secretion (Corrigan et al. Endocrinology, 128:1682-1684 (1991)), and that FS acts similarly by binding to activin, thereby neutralizing its FSH-releasing activity (Kowaga et al., Endocrinology 128, 1682-1684 (1991)).